Table 1.
Characteristics of the study population.
Group differences were tested using t-tests and one-way ANOVAs. Wilcoxon or Kruskall Wallis tests were used to compare non-parametric distributions between two or more groups to compare group differences.
Fig 1.
PON2 protein levels in human bronchial epithelial cells from obese and non-obese asthmatics and control subjects.
Human bronchial epithelial cells from obese-asthmatics subjects had the lowest PON2 levels when compared with healthy controls. Densitometry of PON2 *p<0.05, N = 4/group one-way ANOVA.
Fig 2.
PON2 gene expression exhibits similar trends to the PON2 protein concentration levels measured using Western blot, with a trend towards decreased PON expression in asthma-obese (n = 5, 11.6 ± 0.22; mean ± standard error in VST-normalized messenger RNA levels) compared to lean participants (n = 4, 11.9 ± 0.062).
The error bars depicit mean ± standard error.
Fig 3.
Maximum projection of confocal images stacks (left) of primary HBECs labeled with PON2 (green) and COX IV (red) antibodies and stained with DAPI to mark nuclei. PON2 localizes to mitochondria in primary human bronchial epithelial cells (a), Quercetin treatment does not shift PON2 mitochondria localization and increases its expression in human bronchial epithelial cells (b). Relative intensity of immunolabeling images showed that quercetin treatment on HBEC cultures increases approximately 50% PON2 protein levels (c).
Fig 4.
Ciliated, mucus-producing primary airway epithelial cells, maintained for 21 days in air-liquid interface (ALI) were treated with 25 μM or 50μM of quercetin.
25 μM quercetin significantly increased PON2 protein levels compared to control. Densitometry of PON2 *p<0.005, N = 3/group one-way ANOVA.
Fig 5.
Quercetin reduces cellular and mitochondrial oxidative stress in human airway epithelial cells.
Human bronchial epithelial cells in ALI state, control, and treated with 25μM quercetin. A) H2O2 production p<0.0001, N = 4. B) Mitochondrial ROS production p<0.05, N = 5 one-way ANOVA.
Fig 6.
Quercetin cytotoxicity: Human bronchial epithelial cells in ALI state untreated (Control), treated with 25μM quercetin and positive control (Triton X-100, 0.2%), N = 9/group one-way ANOVA.
Fig 7.
Paraoxonase 2(PON2) knockdown Human Bronchial Epithelial cells (BEAS-2B).
At 48h post-transfection, the expression of PON2 were significantly decreased in PON2 knockdown cells. Relative quantification values were determined relative to scrambled siRNA-treated controls cells and ratio of PON2: GAPDH bands was calculated (N = 6) p<0.0009 Unpaired t test.
Fig 8.
PON2 knockdown rendered cells more susceptible to oxidative stress.
The oxidative stress response in siRNA transfected BEAS2B cells treated with quercetin. BEAS2B transfected cells were pretreated for 24 hs with 25μM quercetin. After washout, cells were exposed for 20 minutes to 25μM DMNQ. H2O2 production was measured by Amplex Red p<0.0007, N = 5/group.one-way ANOVA.